Lecanicillium primulinum,a new hyphomycete (Cordycipitaceae) from soils in the Okinawa's main island and the Bonin Islands,Japan

Lecanicillium primulinum, a new hyphomycete classified in the Cordycipitaceae (Hypocreales, Ascomycota), was isolated from soils collected from the Okinawa's main island and the Bonin Islands, Japan. Using a combination of micro-morphological characteristics and sequences of the ribosomal RNA genes and ITS regions, the isolates were identified as a species of Lecanicillium which was previously undescribed. Phylogenetically the species is close to L. acerosum and Lecanicillium sp. 1 and morphologically it is similar to L. longisporum, L. psalliotae and Lecanicillium sp. 1, but its microconidia differ and it is clearly separated phylogenetically.     

Lecanicillium and Verticillium species from Indonesia and Japan including three new species

Forty-six Lecanicillium strains and one Verticillium strain were isolated from subterranean and epiphytic arthropods, soil, and other sources collected in Indonesia and Japan. These strains were identified as nine Lecanicillium and one Verticillium species including six undescribed species based on light microscopy and the sequences of the ITS-1 and ITS-2 regions including 5.8S ribosomal DNA. Four of the ten species (L. araneicola, L. kalimantanense, Lecanicillium sp. 4, and V. indonesiacum) were recovered from Indonesia, five of the ten (L. attenuatum, L. fusisporum, L. psalliotae, Lecanicillium sp. 1, and Lecanicillium sp. 3) were from Japan, and L. saksenae was from both countries. In this article, new species (L. araneicola, L. kalimantanense, and V. indonesiacum) and a new combination (L. saksenae) are proposed from the fungi isolated from epiphytic and subterranean arthropods collected in East Kalimantan.     

The survival of micromycetes exposed to space conditions

Original data on the survival of fungal spores exposed to space conditions are presented. The experiment was carried out on the Earth-orbiting Russian satellite Foton-M4. The flight duration of the satellite was 45 days. Thirteen fungal species (hyaline as well as pigmented) from 10 genera recovered from destructed stone materials were studied. Sterile quartz sand was inoculated by the fungal spores and was placed into Eppendorf tubes. During the space flight, the Eppendorf tubes with fungal spores were kept inside the Foton descent capsule in the “Biokont” containers and on the external surface of the capsule in the “Exobiofrost” containers exposed to the open space as well. Spores of ten species (77% of all tested species), i.e. Acremonium charticola, Aspergillus niger, Aspergillus versicolor, Chaetomium globosum, Cladosporium sphaerospermum, Penicillium chrysogenum, Penicillium verrucosum, Purpureocillium lilacinum, Sarocladium kiliense, and Trichoderma harzianum, survived after the flight both inside and outside the descent capsule. Only three species (23% of all tested species), i.e. Acremonium furcatum, Engyodontium album and Verticillium zaregamsianum, failed to survive outside as well as inside the capsule. Spore viability differed depending on the fungal species. Thus, spores of some fungal species are able to survive under the complex of stress factors such as low temperature values, radiation, etc. We have shown that micromycetes can be used as a model group for study of eukaryotic organisms’ resistance to stress factors, due to their high tolerance not only to extreme terrestrial environments, but to the extraterrestrial ones as well.     

Effect of inhibitors on calcium carbonate deposition mediated by freshwater algae

Nine green algae, a diatom and three cyanobacteria were shown to precipitate CaCO₃ in batch culture, when grown in the light in a hard water medium containing 68 mg L⁻¹ soluble calcium. The composition of the medium was based on that found in a natural hardwater marina where precipitation of CaCO₃ within algal biofilms occurred. Deposition occurred as a direct result of photosynthesis which caused an increase in the pH of the medium. Once a critical pH had been reached, typically approximately pH 9.0, precipitation began evidenced by a fall in the concentration of soluble calcium in the medium. Certain characteristics of the precipitation process displayed by the diatom Navicula sp. were different to those of the other algae. All algae produced extracellular crystals of irregular morphology. Using a standardized protocol employing the green algae Chlorococcum sp. and Stigeoclonium variabile, the effects of various inhibitors of CaCO₃ nucleation or growth of crystals were studied. Fifteen compounds were screened and assessed for their performance in this context. Most materials effectively delayed deposition of CaCO₃, many decreased precipitation rates and all had a marked effect on crystal morphology. The most effective compound was HEDP (1-hydroxyethylene 1,1 diphosphonic acid), which inhibited precipitation completely at a concentration of 2.5 mg L⁻¹ The use of such compounds to reduce the precipitation of calcium salts within algal biofilms in natural hard waters is discussed.     

Potential of Lecanicillium species for dual microbial control of aphids and the cucumber powdery mildew fungus, Sphaerotheca fuliginea

Detached leaf disc bioassays were conducted against cucumber powdery mildew and three species of aphid with three entomopathogenic species of Lecanicillium; Lecanicillium longisporum (Vertalec^®), Lecanicillium attenuatum (CS625), and an unidentified isolate (DAOM198499). The three Lecanicillium species had high virulence against the aphids Myzus persicae, Macrosiphum euphorbiae and Aulacorthum solani with the exception of DAOM 198499, which demonstrated reduced virulence to A. solani with an LT₅₀ of 6.4 days. Otherwise, LT₅₀ ranged between two and four days. Suspensions of conidia and blastospores of the Lecanicillium species were also applied onto 15 mm leaf discs dissected from cucumber plants previously inoculated with Sphaerotheca fuliginea. Powdery mildew did not develop when the Lecanicillium applications were made one and eight days after S. fuliginea inoculations. When Lecanicillium was applied to highly infected leaf discs 11 and 15 days after S. fuliginea inoculation, the application suppressed subsequent production of S. fuliginea spores as compared to the controls. These results suggest the potential of a dual role for Lecanicillium spp. as microbial control agents against aphids and powdery mildew.     

Functions of Calcium-bound Phosphorus in Relation to Characteristics of Phosphorus Releasing Bacteria in Sediment of a Chinese Shallow Lake (Lake Wabu)

Sediment phosphorus (P) release accelerates lake eutrophication, while retention capacity and release potential of different P fractions, calcium-bound P (CaCO₃～P) in particular, still remains unclear. Fractionation and sorption behaviors of phosphorus were studied in sediment of a Chinese shallow lake (Lake Wabu) and two inflowing rivers from December 2011 to December 2012. Abundance of P releasing bacteria was analyzed, and their main species were isolated using a culture-dependent method and identified by their 16S rDNA sequences. CaCO₃～P release abilities of these bacteria were also tested. In sediments of both the lake and rivers studied, the rank order of the different P extracts was CaCO₃～P > iron-bound P > acid-soluble organic P > hot NaOH-extractable organic P. At the same time, CaCO₃～P content and equilibrium P concentration (EPC₀) values in river sediments were significantly higher than those in the lake. Additionally, EPC₀ changes non-monotonically with increasing CaCO₃～P content, forming a V-shaped curve, with the lowest EPC₀ at an intermediate CaCO₃～P content (around 180 mg kg^?1). Below this threshold, CaCO₃～P was a component strengthening P retention; moreover, CaCO₃～P became an active species responsible for P release. Noticeably, between the two parts divided by this threshold, the differences in abundance of inorganic phosphorus solubilizing bacteria (IPB) and organic phosphorus mineralizing bacteria (OPB) were insignificant and the dominant IPB species clustered together. By contrast, OPB was distinguished from each other, whose dominant species isolated from the part with higher CaCO₃～P content, namely Novosphingobium sp., exhibited a stronger ability to solubilize CaCO₃～P. Shortly, with lower content, CaCO₃～P tends to stabilize P in sediment; while with higher content or under eutrophic condition, it shifted into P source, with some OPB species becoming the main factors to drive its release.     

Potential of Lecanicillium spp. for management of insects, nematodes and plant diseases

Fungi in the genus Lecanicillium (formerly classified as the single species Verticillium lecanii) are important pathogens of insects and some have been developed as commercial biopesticides. Some isolates are also active against phytoparasitic nematodes or fungi. Lecanicillium spp. use both mechanical forces and hydrolytic enzymes to directly penetrate the insect integument and the cell wall of the fungal plant pathogen. In addition to mycoparasitism of the plant pathogen, the mode of action is linked to colonization of host plant tissues, triggering an induced systemic resistance. Recently it was demonstrated that development of Lecanicillium hybrids through protoplast fusion may result in strains that inherit parental attributes, thereby allowing development of hybrid strains with broader host range and other increased benefits, such as increased viability. Such hybrids have demonstrated increased virulence against aphids, whiteflies and the soybean cyst nematode. Three naturally occurring species of Lecanicillium, L. attenuatum, L. longisporum, and an isolate that could not be linked to any presently described species based on rDNA sequences have been shown to have potential to control aphids as well as suppress the growth and spore production of Sphaerotheca fuliginea, the causal agent of cucumber powdery mildew. These results suggest that strains of Lecanicillium spp. may have potential for development as a single microbial control agent effective against several plant diseases, pest insects and plant parasitic nematodes due to its antagonistic, parasitic and disease resistance inducing characteristics. However, to our knowledge, no Lecanicillium spp. have been developed for control of phytopathogens or phytoparasitic nematodes.     

Organic Acid Production by Basidiomycetes: I. Screening of Acid-Producing Strains

Sixty-seven strains belonging to 47 species of Basidiomycetes were examined for their acid-producing abilities in glucose media, in both the presence and absence of CaCO₃, in stationary and shake cultures. Some strains were found to produce large quantities of oxalic acid. The oxalic acid-producing strains could be separated into two groups. Strains of one group (mostly brown-rot fungi) were able to produce oxalic acid, regardless of whether CaCO₃ was present in the medium. Strains of the other group (mostly white-rot fungi) were characterized by their ability to produce oxalic acid only when CaCO₃ was added to the medium. With the latter group, shake-culturing was generally more effective than stationary culturing in respect to acid production. In the CaCO₃-containing media, Schizophyllum commune, Merulius tremellosus, and Porodisculus pendulus were found to produce substantial amounts of L-malic acid as a main metabolic product, along with small quantities of oxalic and other acids in shake cultures. Especially, S. commune and M. tremellosus may be employed as malic acid-producing species.     

Microfungal Contaminants on Mobile Phones of Health Services Vocational School Students in Marmaris,Turkey

In this study, it was aimed to determine microfungi on mobile phones. Totally, 50 mobile phones were used belonging to Health Services Vocational School students. The samples were taken by swabbing the screen and keys of mobile phones using moistened sterile swab sticks. A total of 24 different microfungal species were obtained belonging to Alternaria, Aspergillus, Cladosporium, Geotrichum, Penicillium, Phoma, Rhinocladiella, Scopulariopsis, Trichoderma, and Trichophyton genera. The genera of microfungi most abundant in terms of the number of species on the mobile phones were Aspergillus, Cladosporium, and Penicillium. Numerically, Cladosporium was found as the most abundant on the mobile phones. Cladosporium herbarum colonies were highest in number, followed by Cladosporium sphaerospermum, and Penicillium verrucosum var. cyclopium. When percentages of each species present on the mobile phones were considered, C. herbarum and C. sphaerospermum were the most common. There was a great similarity between the dominant microfungi isolated from mobile phones and dominant microfungi obtained from studies of atmospheric microfungi in Turkey.     

Production of Concentrated Lactococcus lactis subsp. cremoris Suspensions in Calcium Alginate Beads

The effect of simultaneous modification of medium composition and growth conditions on the production of Lactococcus lactis subsp. cremoris biomass in calcium alginate beads was studied by the response surface method. Statistical methods of data analysis for unbalanced experiments are illustrated. The media tested were whey, whey supplemented with yeast extract and/or meat extract, milk, and the commercial medium Gold Complete (Nordica). Fermentations were performed at 23°C under pH control (5.6, 6.0, 6.4, or 6.8). In one complete series, 1% CaCO₃ was added to the growth media. There were strong interactions between CaCO₃ and media, CaCO₃ and pH level, and CaCO₃, media, and pH level. In media with CaCO₃, all first-order interactions between media, pH, and sampling time were significant. The addition of CaCO₃ increased cell counts in whey-meat extract medium, but no significant difference was found with the other media. Uncoupling between growth and acidification occurred between 16 and 22 h. Highest counts were obtained on milk and Gold Complete (6 × 10¹⁰/g). In CaCO₃-containing media, pH influenced cell counts only in whey and in Gold Complete (pH 5.6 and 6.0 giving the best results); pH also influenced the bead mass obtained at the end of the fermentation. Biomass production in alginate gels is proposed as a method of obtaining concentrated cell suspensions without centrifugation or filtration.     

Highly thermostable carbonic anhydrase from Persephonella marina EX-H1: Its expression and characterization for CO2-sequestration applications

The codon-optimized carbonic anhydrase gene of Persephonella marina EX-H1 (PMCA) was expressed and characterized. The gene with the signal peptide removed, PMCA(sp−), resulted in the production of approximately five times more purified protein than from the intact gene PMCA using an Escherichia coli expression system. PMCA(sp−) is formed as homo-dimer complex. PMCA(sp−) has a wide pH tolerance (optimum pH 7.5) and a high thermostability even at 100 °C (88 min of thermal deactivation half-life). The melting temperature for PMCA(sp−) was 84.5 °C. The apparent k_cat and K_m values for CO₂ hydration were 3.2 × 10⁵ s⁻¹ and 10.8 mM. The activity of the PMCA(sp−) enzyme was enhanced by Zn²⁺, Co²⁺, and Mg²⁺, but was strongly inhibited by Cu²⁺, Fe³⁺, Al³⁺, Pb²⁺, Ag⁺, and Hg²⁺. PMCA(sp−) readily catalyzed the hydration of CO₂, precipitating CaCO₃ as calcite in the presence of Ca²⁺.     

Importance of B4 Medium in Determining Organomineralization Potential of Bacterial Environmental Isolates

B4 precipitation medium has been used as the preferred medium for studying mineral precipitation using bacterial strains in vitro since pioneer studies were performed by Boquet and coworkers in 1973. Using this medium, several authors have demonstrated that some environmental isolates were able to precipitate minerals, yet others did not. The main goal of the current study is to understand whether pH and buffer conditions would have a significant effect on mineral precipitation results for environmental isolates grown on B4. For this study, a total of 49 strains isolated from natural environments from Puerto Rico were grown on B4 plates, and their CaCO₃ precipitation potential was investigated. Our findings revealed a strong correlation between a lack of CaCO₃ precipitation and the acidification of the B4 plates by the colonies. The ability to precipitate CaCO₃ could be restored by buffering the B4 medium to a pH of 8.2. Buffering capacity of the medium was proposed to be involved in CaCO₃ precipitation: acid-base titrations conducted on the individual ingredients of B4 showed that yeast extract has a poor buffering capacity between pH 6.5–7.5. This pH range corresponds to the pH of B4 plates [6.87 (±0.05)] prior to the inoculation. This might explain why B4 is such a good precipitation medium: a small variation in the H⁺/OH^? balance during microbial growth and precipitation produces rapid changes in the pH of the medium. Finally, an amorphous matrix was distributed within 90% of the examined crystals generated on B4 medium by the environmental strains. Supplemental materials are available for this article. Go to the publisher's online edition of Geomicrobiology Journal to view the free supplemental file.     

Rock phosphate solubilization by four yeast strains

The solubilization of rock phosphate (RP) by four yeast strains, Rhodotorula sp., Candida rugosa, Saccharomyces cerevisiae and Saccharomyces rouxii, which were isolated from wheat rhizospheric soils, was investigated in this study. The yeast isolates demonstrated diverse levels of soluble phosphate releasing abilities in modified Pikovskaya liquid medium containing RP as sole phosphate source. C. rugosa was the most effective solubilizer under different conditions, followed by Rhodotorula sp., S. rouxii and S. cerevisiae. Acidification of the broth seemed to be the major mechanism for RP solubilization by the yeast isolates, and the increase in soluble phosphate released was correlated significantly with an increase in titratable acidity and a drop in pH. The optimal composition for the solubilization of RP by the yeast isolates in the broth was 20 g L^?1 glucose, 1 g L^?1 yeast extract, 0.5 g L^?1 (NH₄)₂SO₄, and 5 g L^?1 RP, respectively. The yeast isolates were able to solubilize RP at wide range of temperature and initial pH, with the maximum percentage of soluble phosphate released being recorded at 30–35 °C and pH 5–6, respectively.     

Effects of neutralizing agents on lactic acid production by Rhizopus oryzae using sweet potato starch

A neutralizing agent is usually employed to counteract the pH reduction during lactic acid fermentation by Rhizopus oryzae. Calcium carbonate (CaCO₃) is used as such a pH controlling agent. The low solubility of CaCO₃ in the fermentation broth could however lead to low efficiency in pH control and cause problems in the subsequent purification process. Therefore, an alternative agent in place of CaCO₃ was examined in this study. The effect of four different neutralizing agents, including CaCO₃, sodium hydroxide (NaOH), ammoniacal solution and sodium bicarbonate (NaHCO₃) on lactic acid production and the morphology of the pellets were investigated. Results indicated that CaCO₃ was still the preferred choice, because of the pellet morphology and the highest lactic acid concentration (43.3 g/L) obtained in the batch using 60 g/L of sweet potato starch as feedstock. It is noteworthy that the lactic acid purification is relatively easier when using NaHCO₃ instead of CaCO₃, due to the higher solubility of sodium lactate than calcium lactate. Therefore, even the batch with CaCO₃ had a slightly higher productivity (1.23 g/L/h) than the batch with NaHCO₃ (1.14 g/L/h), NaHCO₃ might be the first choice for process designers whenever recovery is vital.     

Monosaccharide-Responsive Phenylboronate-Polyol Cell Scaffolds for Cell Sheet and Tissue Engineering Applications

Analyte-responsive smart polymeric materials are of great interest and have been actively investigated in the field of regenerative medicine. Phenylboronate containing copolymers form gels with polyols under alkaline conditions. Monosaccharides, by virtue of their higher affinity towards boronate, can displace polyols and solubilize such gels. In the present study, we investigate the possibility of utilizing phenylboronate-polyol interactions at physiological pH in order to develop monosaccharide-responsive degradable scaffold materials for systems dealing with cells and tissues. Amine assisted phenylboronate-polyol interactions were employed to develop novel hydrogel and cryogel scaffolds at neutral pH. The scaffolds displayed monosaccharide inducible gel-sol phase transformability. In vitro cell culture studies demonstrated the ability of scaffolds to support cell adhesion, viability and proliferation. Fructose induced gel degradation is used to recover cells cultured on the hydrogels. The cryogels displayed open macroporous structure and superior mechanical properties. These novel phase transformable phenylboronate-polyol based scaffolds displayed a great potential for various cell sheet and tissue engineering applications. Their monosaccharide responsiveness at physiological pH is very useful and can be utilized in the fields of cell immobilization, spheroid culture, saccharide recognition and analyte-responsive drug delivery.     

Soil preference of populations of genotypes ofAsplenium trichomanes L. andPolypodium vulgare L. in Belgium as related to cation exchange capacity

A number of populations ofPolypodium vulgare L. andAsplenium trichomanes L. were sampled with corresponding soils in Belgium in order to get an idea of their suitability for the investigation of related calcicole and calcifuge taxa. Morphology and cytology enabled us to distinguish the subspecies and the hybrids. Analyses of the soils for pH, CaCO₃, Al and H show that subspecies and hybrids have distinct soil preferences and can be characterised as calcicoles-neutrophile/basiphiles or calcifuge-acidiphiles. Physiological implications of the ecological status of the taxa are discussed in the light of their root cation exchange capacity.     

Template preferences of DNA polymerase and nuclease activities appearing during early development of Xenopus laevis.

The three DNA polymerase activities identified in early embryos of Xenopus laevis(Benbow et al., 1975) were further distinguished by their template preferences, pH optima and sensitivity to monovalent cations. In addition, endo and exonuclease activities at pH 7.5 were identified and monitored through early development. Endonuclease activity increased throughout early development while exonuclease activity remained nearly constant. These nuclease activities were not responsible for the different template preferences of the partially purified DNA polymerase activities.     

Observations on the proventricular glands (‘organes racémiformes’) of the oribatid miteChamobates borealis (Acari,Oribatida): an organ of interest for studies on adaptation of animals to acid soils

The proventricular glands of the oribatid miteChamobates borealis (Trägårdh, 1902) were investigated by electron microscopy and histochemistry, and their function was tested in a laboratory experiment. Specimens of the same species collected during a field study also were investigated.The cells of the proventricular glands are characterized by great amounts of mineral spherites and seem to be sensitive to alterations of environmental pH and concentration of CaCO₃. It was demonstrated that a reduction of pH leads to a decrease of the spherites, whereas at a high pH the number of spherites increases.The proventricular glands with their spherites seem to play an important role in regulation of mineral budget, pH and detoxification of heavy metals.